Biochemical characterization of skin fibroblasts derived from WHHL-rabbit, a notable animal model for familial hypercholesterolemia.

Abstract

WHHL-rabbit (Watanabe-heritable hyperlipidemic rabbit), a strain of animal with spontaneous hyperlipidemia, is characterized by the remarkable resemblance of the pathology to type II hyperlipoproteinemia in man. In order to understand the biological basis of the disorder in the WHHL-rabbit, the lipid metabolism of cultured skin fibroblasts from the mutant rabbits was studied. Skin fibroblasts from WHHL-rabbits grown in the presence of whole serum showed 12.3-times and 3.5-times higher activities of sterol and fatty acid syntheses from [14C]acetate, respectively, when compared with fibroblasts from normal rabbits. The cholesterol esterification activity, which was measured by the incorporation of [14C]oleate into cholesterol esters, however, was 9-times lower in the mutant cells than the normal cells. The higher sterol-synthesizing capacity of the WHHL-rabbit fibroblasts was attributed to the 5-times-enhanced activity of 3-hydroxy-3-methylglutaryl-CoA reductase, the rate-limiting enzyme in cholesterol synthesis, compared with that in normal cells. In normal cells, the removal of lipoproteins from the culture medium caused a marked enhancement of 3-hydroxy-3-methylglutaryl-CoA reductase activity with a concomitant decrease of cholesterol esterification activity. The incubation of normal cells with low density lipoprotein (LDL) led to a reciprocal decrease in 3-hydroxy-3-methylglutaryl-CoA reductase activity and increase in the esterification activity. In contrast, the high level of 3-hydroxy-3-methylglutaryl-CoA reductase, which was associated with low cholesterol-esterification activity, in the WHHL-rabbit cells was essentially unchanged by removal or addition of LDL, indicating the absence of the functional LDL-mediated regulation system in the mutant cells. The studies using 125I-labeled LDL showed that normal cells possessed the specific receptor for LDL; LDL was bound, internalized by the receptor and degraded within cells. On the other hand, the finding that only 6 % of the receptor activity was present in the mutant cells when compared with normal cells, revealed that the mutant cells were the receptor-defective type. Thus, the primary defect in WHHL-rabbit fibroblasts is the lack of LDL receptor which regulates both 3-hydroxy-3-methylglutaryl-CoA reductase and cholesterol esterification activities. The mode of inheritance of LDL receptor-deficiency in WHHL-rabbit cells was found to be recessive, since all cell lines from the heterozygous rabbits possessed almost the same LDL receptor activities as normal cells. WHHL-rabbit, the first LDL receptor-deficient animal. can be a suitable model for familial hypercholesterolemia in man.